The process of lymphocyte development and differentiation is tightly controlled by cytokines. B lymphocyte stimulating factors (BLyS, also called BAFF, TNSF13B, THANK, TALL-1, and zTNF4) and a proliferation-inducing ligand (APRIL) belong to the tumor necrosis factor (TNF) and tumor necrosis factor receptor (TNFR) superfamilies that play a critical role in the survival and death of immune cells (Schneider, 2005, Current opinion in immunology, 17:282-289; Seyler et al., 2005, The Journal of Clinical investigation, 115:3083-3092). They provide key signals for the development and proliferation of B lymphocytes and T lymphocytes. In addition, they suppress lymphocyte apoptosis and stimulate the production of immunoglobulins in the blood (Wang et al., 2001, Nature Immunology, 2:632-637).
The over-expression of BLyS and APRIL, however, are associated with many lymphoproliferative disorders, as well as immunodeficiency and autoimmune diseases. These conditions include, for example, chronic lymphocyte leukemia, multiple myeloma, B lymphocyte lymphoma, systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), and sicca syndrome. Further, clinical studies have shown that the degree of BLyS and APRIL over-expression correlates to the severity of diseases.
BLyS and APRIL regulate immune responses through binding to their receptors on the cell surface. Their pro-survival function is mainly regulated through the binding of BLyS to BAFF-R and B cell maturation antigen (BCMA) and/or APRIL to BCMA.
In contrast, transmembrane activator and calcium modulator and cyclophilin ligand interactor (TACI) function at least in part as a negative regulator of BLyS and APRIL activity. TACI has an extracellular domain, a transmembrane domain and an intracellular domain, wherein the extracellular domain includes an amino terminal region, a cystein-rich region and a stalk region. Because the cystein-rich region has high affinity to both BLyS and APRIL, the expression of TACT inhibits the pro-survival signaling pathway by blocking the binding of BLyS and APRIL to BAFF-R and/or BCMA.
Pre-clinical studies have shown that TACI or TACI-fusion proteins are useful for treating or ameliorating pathological immune conditions such as lymphoproliferative disorders, and autoimmune diseases.
Proprotein convertase (PC) is a group of enzymes that cleave proteins at specific sites. For many secretory protein or peptides, PC cleavage is a part of normal protein post-translational modification process for protein maturation or activation. For other proteins or polypeptides, the existing of PC cleavage sites within protein sequences may cause undesired protein cleavage. When the TACI sequence is analyzed by using the proprotein convertase (PC) artificial neural network computer program, a number of PC cleavage sites can be identified. PC cleavage at various sites results in TACI proteins that are heterogeneous in size, structure, stability, and activity. This enzymatic cleavage process also generates unwanted protein products.
To address this problem, PCT/US2002/015910 provides a method for avoiding PC cleavage by constructing a TACI-Fc fusion protein having no amino terminal region and only part of the stalk region. However, it is reported that the TACI-Fc fusion protein with no amino terminal region exhibits significantly reduced biological activity (Wu et al., 2000, The Journal of Biological Chemistry, 275:35478-35485). Consequently, there remains a substantial need for improved TACI-Fc fusion proteins that not only possess high binding affinity to BLyS and/or APRIL, but also exhibit biological activity similar to the native TACI protein. In addition, a need exists for improved methods of recombinant production of TACI proteins that are capable of yielding homogeneous biologically-active TACI protein products with lot-to-lot consistency.